Albright hereditary osteodystrophy (AHO) is an autosomal dominant disorder characterized by short stature, obesity, subcutaneous ossifications, and brachydactyly. Some family members have AHO features only (pseudopseudohypoparathyroidism, PPHP) while others have AHO in association with resistance to multiple hormones which activate Gs-coupled receptors (pseudohypoparathyroidism type Ia, PHP Ia). AHO/PHPIa is caused by heterozygous Gs-alpha null mutations and most affected patients have a 50% deficiency in Gs-alpha subunit function and/or expression in peripheral tissues (both PHP Ia and PPHP). Gs-alpha is a ubiquitously expressed G protein alpha-subunit that is required for the cyclic AMP response to hormones and other extracellular signals. We have shown in mice and more recently in humans that Gs-alpha is imprinted in a tissue-specific manner. In some hormone target tissues Gs-alpha is expressed primarily from the maternal allele, and therefore maternal inheritance of Gs-alpha mutations results in PHP Ia while paternal inheritance of these same mutations leads to PPHP. More recently GNAS/Gnas has been shown to be a very complicated gene with multiple imprinted gene products generated by several alternative promoters and first exons. NESP55 is a chromogranin-like protein that is maternally expressed while XL-alpha-s is a paternally expressed Gs-alpha isoform with a long amino-terminal extension. Both are primarily expressed in neuroendocrine tissues. We have shown that NESP imprinting is not established until postimplantation development. Just upstream of the XL-alpha-s promoter is the promoter for a paternally-expressed antisense transcript (NESPAS) that traverses the NESP promoter from the opposite direction. We identified another alternative first exon (exon 1A) that generates paternally expressed untranslated mRNAs and that is a maternal germline imprint mark. We have shown that this region has allele-specific differences in histone methylation. We also have shown that the Gsa promoter and first exon also has allele-specific differences in histone methylation which correlates to its tissue-specific imprinting, even though this region does not undergo DNA methylation. We have shown that PHP Ib (parathyroid hormone resistance in the absence of AHO) is virtually always associated with loss of maternal exon 1A imprinting. A detailed analysis of GNAS imprinting in PHP Ib patients showed that familial cases tend to only have abnormal exon 1A imprinting associated with a deletion mutation within a closely-linked gene, while sporadic cases often have additional imprinting defects involving NESP and XL-alpha-s. In some patients, the imprinting of the XL-alpha-s promoter and its first exon is discordant. We have examined Gnas methylation in mice which do not establish maternal germline imprints (dnmt3L-/-) and show that the whole Gnas locus develops a paternal methylation pattern on both alleles, indicating that imprinting of the whole locus depends on maternal germline imprints. We have also generated exon 1A knockout mice, and show that this region is not required for Nesp and XL-alpha-s imprinting, but is required for tissue-specific Gs-alpha imprinting. Mice with paternal exon 1A deletion, which have Gs-alpha overexpression in renal proximal tubules, have increased parathyroid hormone sensitivity with low circulating parathyroid hormone levels. We have also examined the effects of Gs-alpha deficiency in chondrocytes and osteoblasts in mice. Loss of Gs-alpha in chondrocytes leads to a growth plate phenotytpe similar to loss of PTHrP or its receptor, and reminiscent of the growth plate lesion in AHO. Osteoblast-specific deficiency leads to a low bone turnover state with differential effects in cortical and trabecular bone.